Analysis fo the role of a novel ovarian protein during oocyte maturation using genetic engineering procedures

从猪卵巢卵泡液中分离到一种80 kDa的蛋白水解酶--卵泡素。该酶能降解合成肽和天然肽底物的Arg-X键，并与人血浆激肽释放酶高度同源。该酶对抑肽酶、苯甲酰胺、亮肽酶、止痛剂和氟磷酸二异丙酯有较强的抑制作用，表明该酶为丝氨酸蛋白酶。研究发现，人的单链组织型纤溶酶原激活物被该酶有效地转化为双链形式。这一发现表明，丝氨酸蛋白酶参与了蛋白水解级联纤溶酶原激活物/纤溶酶系统的激活，该系统通常被认为在排卵过程中发生卵泡破裂。猪前卵泡素是从卵泡液中分离出来的。经胰酶处理后，它很容易被激活。研究还表明，猪卵巢卵泡液中含有一种促卵泡素激活酶的活性。在体外受精过程中从人卵巢获得的卵泡液中含有一种与合成的含Arg内肽酶底物检测到的猪卵泡素类似的酶。纯化和性质研究表明，该酶为血浆激肽释放酶。血浆激肽释放酶与蛋白水解酶抑制剂α2-巨球蛋白的结合被证明是该酶在液体中的主要形式，表明该酶的活性受该抑制物的调节。研究还表明，卵泡液中存在的血浆激肽释放酶可能来自肝脏。

A 80kDa proteinase, follipsin, was isolated from the follicular fluid of porcine ovaries. The enzyme hydrolyzed internal Arg-X bonds of synthetic and natural peptide substrates, and was highly homologous wiht human plasma kallikrein. The enzyme was strongly inhibited by aprotinin, benzamidine, leupeptin, antipain, and diisopropylfluorophosphate, indicating that it is a serine proteinase. It was found that human single-chain tissue-type plasminogen activator was efficiently converted to its two-chain form by the enzyme. The finding suggests the involvement of the serine proteinase in triggering the activation of proteolytic cascade plasminogen activator/plasmin system, that is generally thought to operate in follicle rupture during ovulation.Porcine profollipsin was isolated from the fluid. It was readily activated by trypsin treatment. The study also showed that the follicular fluid of porcine ovary contains a profollipsin-activating enzyme activity.Follicular fluid from human ovaries obtained in in vitro fertilization procedure contained an enzyme similar to porcine follipsin as detected with synthetic, Arg-containing endopeptidase substrates. Purification and characterization studies showed that the enzyme was plasma kallikrein. Plasma kallikrein in association with the proteinase inhibitor alpha 2-macroglobulin was demonstrated to be a predominant form of the enzyme in the fluid, suggesting that the enzyme activity is regulated by the inhibitor. The study also indicated that the plasma kallikrein present in follicular fluid probably comes from the liver.